• 한국동물생명공학회지
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• 제36권4호
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• pp.183-188
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• 2021

In vitro maturation (IVM) of oocytes is the procedure where the immature oocytes are cultivated in a laboratory until they are mature. Since IVM oocytes generally have low developmental competence as compared to those matured in vivo, development of an optimal IVM culture system by fine-tuning culture conditions is crucial to maintain high quality. In-depth knowledge and a deep understanding of the in vivo physiology of oocyte maturation are pre-requisites to accomplish this. Within ovarian follicles, various signaling pathways that drive oocyte development and maturation regulate interaction between oocytes and surrounding somatic cells. This review discusses the sonic hedgehog (SHH) signaling pathway, which has been demonstrated to be intimately involved in folliculogenesis and oocyte maturation. Advances in elucidating the role of the SHH signaling pathway in oocyte maturation will aid attempts to improve the current inferior in vitro oocyte maturation system.

• Reproductive and Developmental Biology
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• 제39권2호
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• pp.37-41
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• 2015

The oocyte undergoes various events during maturation and requires many substances for the maturation process. Various intracellular organelles are also involved in maturation of the oocyte. During the process glucose is essential for nuclear and cytoplasmic maturation, and adenosine triphosphate is needed for reorganization of the organelles and cytoskeleton. If mitochondrial function is lost, several developmental defects in meiotic chromosome segregation and maturation cause fertilization failure. The endoplasmic reticulum, a store for $Ca^{2+}$, releases $Ca^{2+}$ into the cytoplasm in response to various cellular signaling molecules. This event stimulates secretion of hormones, growth factors and antioxidants in oocyte during maturation. Also, oocyte nuclear maturation is stimulated by growth factors such as epidermal growth factor. This review summarizes roles of organelles with focus on the Golgi apparatus during maturation in oocyte.

• 한국동물생명공학회지
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• 제35권1호
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• pp.2-11
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• 2020

Assisted reproductive technologies (ART) merely depend on improving the oocyte maturation and their developmental competence to produce good quality embryos. Oocyte maturation passes through long and complex molecular steps starts from the early embryonic life and ends with sperm fertilization. Oocyte developmental competence can be attained by improving the nuclear and cytoplasmic mechanisms together with some epigenetic maturation. In this review, we highlight the cornerstones of oocyte maturation on both nuclear and cytoplasmic levels. Interfering or supporting these molecular mechanisms would help in the development of novel regulating agents for reproductive performance of humans and livestock species.

• 한국수정란이식학회지
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• 제33권3호
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• pp.99-105
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• 2018

To determine the differences in the in-vitro ovum maturation process of bovine, we compared the expression of MMPs in these oocytes and cumulus cell throughout oocytes maturated. In an attempt to investigate the effect of MMP activation and inhibitors in total protein of cumulus cell and, oocytes during oocytes maturation, we examined and monitored the localization and expression of MMPs (MMP-2 and MMP-9), TIMPs (TIMP-2 and TIMP-3), as well as their expression profiles (Real-time PCR, Gelatin Zymography and ELISA). Our results that the bovine oocytes MMP-2 and MMP-9 level was significantly associated with the rate of maturity of oocytes (P<0.05). In cumulus cell, MMP-2 was highly expressed in all stages of the oocyte's maturation. The final oocytes maturation exhibited strong gelatinase activity. There was no significant correlation between cumulus cell MMP-9 and the maturation rate of oocytes. However, for the oocyte cytoplasm MMP-9 expression was significant correlation to the maturation oocytes. There was no significant correlation between cumulonimbus cells MMP-9 and oocyte maturation rates; however, for oocyte cytoplasm, MMP-9 expression was significantly correlated with mature oocyte. However, the TIMP-1 and TIMP-2 protein expression patterns are not correlated with the maturation rate of the oocyte. Our results suggest that MMP different expression pattern may regulate the morphological remodeling of oocyte's in the cumulus cell. Further, the MMP-2 expression has a strong relation with a higher maturation rate of the oocyte.

• Animal cells and systems
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• 제5권1호
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• pp.45-50
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• 2001

Rana ovarian follicles consist of oocyte, vitelline envelope, granulosa cells, and theca/epithelial layer. Using scanning electron microscopy, the surface structure of each follicular component was investigated. Changes in oocyte surface during oocyte maturation were also examined. Theca/epithelial layer was almost transparent and some blood vessels and granulosa cells were observed underneath in intact follicle. The number of granulosa cells was estimated to be 6700-7200 per oocyte. The granulosa cells partially overlapped each other and their microvilli penetrated the vitelline membrane via holes present in the vitelline envelope and seemed to be linked to oocyte microvilli. After removal of the vitelline envelope by microforcep, oocyte microvilli were observed on the surface of the devitellined oocyte. The oocyte microvilli formed partial clusters on the surface of white spot area which appears iust before germinal vesicle breakdown (GVBD), whereas they were evenly distributed in other areas. The microvilli became shorter and less dense with oocyte maturation. The lengths of oocyte microvilli in the immature and mature oocyte were 1.5 $\mu$m and 0.6 $\mu$m, respectively. The present study suggests a fundamental structural change occurring on the oocyte surface during maturation.

• Animal cells and systems
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• 제11권1호
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• pp.1-8
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• 2007

Recently, we have shown that some endocrine disruptors, heavy metals, organotins and azoles suppressed steroidogenic enzymes such as P450 side-chain cleavage enzyme (P450scc) and aromatase in bullfrog ovarian follicles. In the present study, by using an amphibian ovarian follicle culture system, we examined the effects of these endocrine disruptors on maturation and ovulation of oocytes from Rana dybowskii in vitro. Ovarian fragments or isolated follicles were cultured for 24 h in a medium containing frog pituitary homogenate (FPH) or progesterone ($P_{4}$) with or without endocrine disruptors, and oocyte maturation (germinal vesicle breakdown, GVBD) and ovulation were examined. Among the organotins, tributyltin (TBT) strongly inhibited both FPH-and $P_{4}-induced$ oocyte maturation ($ED_{50}$:0.6 and 0.7 ${\mu}M$, respectively); however, tetrabutyltin (TTBT) and dibutyltin (DBT) showed only partial suppression, while monobutyltin (MBT) showed no inhibitory effect. All of the organotins suppressed $P_{4}-induced$ oocyte ovulation very effectively at a low concentration, and TBT and DBT exerted an inhibitory effect on FPH-induced ovulation. Among the heavy metals, mercury (Hg), cadmium (Cd) and cobalt (Co) were very effective in inhibiting FPH-induced oocyte maturation and ovulation, while lead (Pb), arsenite (As) and zinc (Zn) were less effective. However, all of the heavy metals suppressed FPH-induced oocyte ovulation at a high dose ($100{\mu}M$). Among the azoles, itraconazole (ICZ), ketoconazole (KCZ) and clotrimazole (CTZ) effectively inhibited FPH-induced oocyte maturation and ovulation, while econazole (ECZ), miconazole (MCZ) and fluconazole (FCZ) were considerably less effective. These results demonstrated that the abovementioned endocrine disruptors exhibited differential effects on oocyte maturation and ovulation in amphibian follicles and that the frog ovarian culture system could be used as an effective experimental tool to screen and evaluate the toxicity of various endocrine disruptors in vitro.

• 대한수의학회지
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• 제36권4호
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• pp.929-939
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• 1996

The present study was undertaken to establish a relationship between bovine follicle size and oocyte diameter, compare the nuclear maturation competence of oocytes of different diameter groups and the nuclear maturation changes in Korean Native Cattle according to in vitro maturation period. To compare the relationship between follicle size and oocyte diameter, follicles were dissected, measured, and assigned to one of the following size categories($4{\geq}mm$, 3-4mm, 2-3mm, 1-2mm, and < 1mm), investigate the maturation competence in the different-sized oocytes, which were divided into three groups( < $110{\mu}m$, 110 - < $120{\mu}m$, and ${\geq}120{\mu}m$). Oocytes were cultured in the culture medium during 0, 6, 12, 18, and 24hrs, respectively, stained, and measured the nuclear maturation degree according to period. When compared the relationship between follicle size and intrafollicular oocyte diameter, oocyte diameters of three groups of ${\geq}3mm$ follicle-sized were significantly higher than < 3mm (p<0.01). After in vitro maturation, the rates reached to MI stage of < $110{\mu}m$ oocyte groups(25%) was higher than $110-120{\mu}m$ and ${\geq}120{\mu}m$ oocyte groups(11 and 10%) reached to the same stage(p<0.01), and the rates throughout MII stage of $110-120{\mu}m$ and ${\geq}120{\mu}m$ and < $110{\mu}m$(70 and 76%) groups were higher than < $110{\mu}m$(35%)(p<0.01). When nuclear maturation rates were measured according to period, < 6hr groups(7 and 10%) showed lower rates reached to MI than ${\geq}12hr$ groups(100%), 24hr groups(76%) revealed higher rates throughout MII than 18hr groups(40%). These results indicate that the preparation of oocyte for the production of in vitro fertilization embryos and nuclear transplantation ones could be adapted, as follicle increased up to appointed size there was a corresponding increase in oocyte diameter, and differences of nuclear maturation rate revealed according to oocyte diameter and maturation period.

• 한국동물생명공학회지
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• 제34권1호
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• pp.10-19
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• 2019

Morphology of cumulus-oocyte-complexes (COCs) at germinal vesicle (GV) stage as one of the evaluation criteria for oocyte maturation quality after in vitro maturation (IVM) plays important roles on the meiotic maturation, fertilization and early embryonic development in pigs. When cumulus cells of COCs are insufficient, which is induced the low oocyte maturation rate by the increasing of reactive oxygen species (ROS) in porcine oocyte during IVM. The ROS are known to generate including superoxide and hydrogen peroxide from electron transport system of mitochondria during oocyte maturation in pigs. To regulate the ROS production, the cumulus cells is secreted the various antioxidant enzymes during IVM of porcine oocyte. Our previous study showed that Mito-TEMPO, superoxide specific scavenger, improves the embryonic developmental competence and blastocyst formation rate by regulating of mitochondria functions in pigs. However, the effects of Mito-TEMPO as a superoxide scavenger to help the anti-oxidant functions from cumulus cells of COCs on meiotic maturation during porcine oocyte IVM has not been reported. Here, we categorized experimental groups into two groups (Grade 1: G1; high cumulus cells and Grade 2: G2; low cumulus cells) by using hemocytometer. The meiotic maturation rate from G2 was significantly (p < 0.05) decreased (G1: $79.9{\pm}3.8%$ vs G2: $57.5{\pm}4.6%$) compared to G1. To investigate the production of mitochondria derived superoxide, we used the mitochondrial superoxide dye, Mito-SOX. Red fluorescence of Mito-SOX detected superoxide was significantly (p < 0.05) increased in COCs of G2 compared with G1. And, we examined expression levels of genes associated with mitochondrial antioxidant such as SOD1, SOD2 and PRDX3 using a RT-PCR in porcine COCs at 44 h of IVM. The mRNA levels of three antioxidant enzymes expression in COCs from G2 were significantly (p < 0.05) lower than COCs of G1. In addition, we investigated the anti-oxidative effects of Mito-TEMPO on meiotic maturation of porcine oocyte from G1 and G2. Meiotic maturation and mRNA levels of antioxidant enzymes were significantly (p < 0.05) recovered in G2 by Mito-TEMPO ($0.1{\mu}M$, MT) treatment (G2: $68.4{\pm}3.2%$ vs G2 + MT: $73.9{\pm}1.4%$). Therefore, our results suggest that reduction of mitochondria derived superoxide by Mito-TEMPO may improves the meiotic maturation in IVM of porcine oocyte.

• Clinical and Experimental Reproductive Medicine
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• 제23권3호
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• pp.357-366
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• 1996

The present experiments aimed to investigate the metabolism of calcium during oocyte maturation in rat. The concentration of free calcium and calmodulin in oocytes was measured respectively by using of fluo-3/AM and FITC with microscope fluorescence spectrometer. The ultrastructural localization of calcium precipitates in oocytes was observed with the transmission electron microscope. Cumulus-free immature oocytes(GV-oocyte) were cultured in vitro through 15 hours. The free calcium concentration in GV oocyte was $55.9{\pm}3.5nM$. In calcium-containing medium, the free calcium concentration was increased in germinal vesicle breakdown(GVBD) oocyte($64.2{\pm}7.3nM$). In normal medium after calcium chelator treatment ($10{\mu}M$ BAPTA/AM), the free calcium contents were slightly lower than those in control group. In calcium-free medium, the free calcium content was drastically increased in GVBD($72.7{\pm}3.4nM$) and metaphase I - anaphase I ($88.0{\pm}3.4nM$) oocyte. In maturation rate of oocytes, GVBD rate was high in control group($82.9{\pm}6.55%$) and calcium chelator treatment group($91.2{\pm}4.4%$), but in calcium-free medium group, it was low and then the oocyte was degenerated without polar body formation. Relative content of calmodulin in oocyte was significantly(P<0.001) increased in metaphase I - anaphase I than in GV and GVBD oocyte. The calcium precipitates were observed in mitochondria and cytoplasm of GV oocyte but that were not observed in mitochondria of GVBD and metaphase I - anaphase I oocyte. And then the calcium precipitates reappeared in mitochondria of metaphase II oocyte. The above results indicate that changes in free calcium and calmodulin concentration of oocyte occur according to the maturational stages and the extracellular calcium is required during oocyte maturation. Also change of calcium localization in oocyte occurs according to the maturational stages.

• Applied Microscopy
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• 제28권3호
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• pp.263-271
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• 1998

Oocyte maturation of the swordtail (Kiphophorus hellerii) was investigated by light and electron microscopy. In the ovary of the swordtail, various staged oocytes were observed, Mature oocytes were located in ovarian cortex, meanwhile immature ones were positioned in ovarian medulla. The oocyte was surrounded by several structures or cells such as chorion, follicle cells, follicular theaca and ovarian epithelium, respectively, from the inside toward outside. Growing and maturing oocytes healed numerous microvilli which interconnected the oocyte and the follicle cells to communicate each other. The mature oocyte had the electron dense chorion which appeared to be ultrastructure of two layers and contained pore canals. Oocyte maturation was characterized by not only the enlarged cell size and well differentiated cell organelles, brit also the increases of fat droplets, pinocytotic vesicles and yolk granules.